Project Summary/Abstract - Project 3: Glioblastoma multiforme (GBM) is the most common malignant brain tumor in the United States but despite the aggressive treatment GBM has a dismal prognosis with a median overall survival of ~15 months. Investigators in Project 3 have found that pharmacological ascorbate (P-AscH-), is a promising adjuvant to chemo-radiation therapy in GBM exploiting fundamental differences in oxidative metabolism. Preliminary data demonstrate the selective toxicity of P-AscH- to GBM cells, relative to normal human astrocytes (NHA), occurs via H2O2-induced oxidative stress mediated by redox active Fe ions. Furthermore, in a pre-clinical GBM model as well as in a phase 1 clinical trial (NCT01752491), they found that P-AscH- (achieving ~20 mM [plasma]) + radiation and temozolomide therapy was well-tolerated with minimal adverse events while showing encouraging therapeutic responses. Finally, preliminary data also support the idea that magnetic resonance (MR) imaging may be capable of detecting the redox state of Fe (i.e., Fe+2/Fe+3) for predicting responses to P-AscH- in combination with chemo-radiation therapy via assessing changes of T2* relaxometry. Currently, there is no clear understanding of mechanisms causing the differential susceptibility of GBM vs. normal brain cells to P-AscH- in vivo. Project 3 will test the hypothesis that P-AscH- selectively increases labile redox active Fe associated H2O2 formation in human GBM cells, relative to normal astrocytes, leading to increased oxidative DNA damage and increased sensitivity of GBM cells to radiation and temozolomide that can be assessed in vivo using MR imaging with T2* relaxometry and quantitative susceptibility mapping. Aim 1 will determine if P-AscH--induced increases in intracellular [H2O2] mediate chemo-radiosensitization in patient derived GBM cells of Proneural, Neural, Classical and Meschymal subtypes relative to normal human astrocytes (NHA) via enhanced DNA double strand breaks and differences in H2O2 metabolism in GBM cells. Aim 2 will determine if increased labile iron pools (LIP) mediate the sensitivity of GBM vs. NHA to P-AscH--induced chemo-radiosensitization. Aim 3 will determine in orthotopic GBM xenograft models if P-AscH- induced regulation of the redox-active LIP and selective sensitization to radiation and chemotherapy can be predicted using T2* MRI imaging and quantitative susceptibility mapping as well as standard biochemical assays. Completion of the aims will define biochemical mechanisms underlying P-AscH--mediated selective chemo-radiosensitization of GBM cells in vitro and in vivo orthotopic models as well as investigating highly innovative MR imaging approaches to predict responses. Project 3 is also well-integrated into the overall theme of the PO1 to provide a new paradigm for exploiting fundamental differences in redox metabolism to improve treatment efficacy in cancer patients using P-AscH-.